Diverse and rapidly evolving viral strategies modulate arthropod vector gene expression

Vector-borne viral diseases threaten human and wildlife worldwide. Vectors are often viewed as a passive syringe injecting the virus, however to survive, replicate and spread, viruses must manipulate vector biology. While most vector-borne viral research focuses on vectors transmitting a single virus, in reality vectors often carry diverse viruses. Yet how viruses affect the vectors remains poorly understood. Here we focused on the varroa mite, an emergent parasite that vectors over 20 honey bee viruses, and has been responsible for colony collapses worldwide, as well as changes in global viral populations. Co-evolution of the varroa and the viral community makes it possible to investigate whether viruses affect vector gene expression, and whether these interactions affect viral epidemiology. Using a large set of available varroa transcriptomes we identified how abundances of individual viruses affect the vector transcriptional network. Perhaps surprisingly, we found no evidence of competition between viruses, but rather that some virus abundances are positively correlated. Furthermore, we found a strong correlation between the extent to which a virus interacts with the vector gene expression and co-occurrence with other viruses, suggesting that interactions with the vector affect epidemiology. We experimentally validated this observation by silencing candidate genes using RNAi and found that reduction in varroa gene expression was accompanied by a change in viral load. Combined, the meta-transcriptomic analysis and experimental results shed light on the mechanism by which viruses interact with each other and with their vector to shape the disease course.

Occurrence and Phylogenetic Analysis of DWV in Stingless Bee (Apidae sp.) in China: A Case Report

Honey bees play a vital role in providing pollination services for agricultural crops and wild flowering plants. However, the spillover risk of their pathogens to other pollinators or wild insects is becoming a cause for concern. There is some evidence that stingless bees can carry honey bee viruses, but little is known about the presence of honey bee viruses in stingless bees in China. Here, we investigate the occurrence of major honey bee pathogens including bacteria, fungi, and viruses in stingless bees (Apidae: sp.). Our results show that the stingless bees (Apidae: sp.) were mainly infected with DWV-A, but no DWV-B and DWV-C. Phylogenetic analysis on fragments of lp, RdRp, and VP3 of DWV-A indicated that genetic variation in VP3 might an important indicator for host-specific viruses, but it requires further study. Our results indicated that DWV-A is not only the major strain of virus currently circulating in managed bee colonies in China and globally, but in stingless bee species as a whole.

An Investigation of Honey Bee Viruses Prevalence in Managed Honey Bees (Apis mellifera and Apis cerana) Undergone Colony Decline

Objective: In the absence of known clinical symptoms, viruses were considered to be the most probable key pathogens of honey bee. Therefore, the aim of this study was to investigate the prevalence and distribution of honey bee viruses in managed Apis mellifera and Apis cerana in China. Methods: We conducted a screening of 8 honey bee viruses on A. mellifera and A. cerana samples collected from 54 apiaries from 13 provinces in China using RT-PCR. Results: We found that the types and numbers of viral species significantly differed between A. mellifera and A. cerana. Black Queen Cell Virus (BQCV), Chronic Bee Paralysis Virus (CBPV), Apis mellifera filamentous virus (AmFV), and Kakugo virus (DWV-A/KV) were the primary viruses found in A. mellifera colonies, whereas Chinese Sacbrood Bee Virus (CSBV) and Sacbrood Bee Virus (SBV) were the primary viruses found in A. cerana. The percentage infection of BQCV and CSBV were 84.6% and 61.6% in all detected samples. We first detected the occurrences of Varroa destructor virus-1 (VDV-1 or DWV-B) and DWV-A/KV in China but not ABPV in both A. mellifera and A. cerana. Conclusion: This study showed that BQCV and CSBV are the major threat to investigated A. mellifera and A. cerana colonies.

Africanized honey bees in Colombia exhibit high prevalence but low level of infestation of Varroa mites and low prevalence of pathogenic viruses

The global spread of the ectoparasitic mite Varroa destructor has promoted the spread and virulence of highly infectious honey bee viruses. This phenomenon is considered the leading cause for the increased number of colony losses experienced by the mite-susceptible European honey bee populations in the Northern hemisphere. Most of the honey bee populations in Central and South America are Africanized honey bees (AHBs), which are considered more resistant to Varroa compared to European honey bees. However, the relationship between Varroa levels and the spread of honey bee viruses in AHBs remains unknown. In this study, we determined Varroa prevalence and infestation levels as well as the prevalence of seven major honey bee viruses in AHBs from three regions of Colombia. We found that although Varroa exhibited high prevalence (92%), its infestation levels were low (4.5%) considering that these populations never received acaricide treatments. We also detected four viruses in the three regions analyzed, but all colonies were asymptomatic, and virus prevalence was considerably lower than those found in other countries with higher rates of mite-associated colony loss (DWV 19.88%, BQCV 17.39%, SBV 23.4%, ABPV 10.56%). Our findings indicate that AHBs possess a natural resistance to Varroa that does not prevent the spread of this parasite among their population, but restrains mite population growth and suppresses the prevalence and pathogenicity of mite-associated viruses.

Intra-Colonial Viral Infections in Western Honey Bees (Apis mellifera)

RNA viruses play a significant role in the current high losses of pollinators. Although many studies have focused on the epidemiology of western honey bee (Apis mellifera) viruses at the colony level, the dynamics of virus infection within colonies remains poorly explored. In this study, the two main variants of the ubiquitous honey bee virus DWV as well as three major honey bee viruses (SBV, ABPV and BQCV) were analyzed from Varroa-destructor-parasitized pupae. More precisely, RT-qPCR was used to quantify and compare virus genome copies across honey bee pupae at the individual and subfamily levels (i.e., patrilines, sharing the same mother queen but with different drones as fathers). Additionally, virus genome copies were compared in cells parasitized by reproducing and non-reproducing mite foundresses to assess the role of this vector. Only DWV was detected in the samples, and the two variants of this virus significantly differed when comparing the sampling period, colonies and patrilines. Moreover, DWV-A and DWV-B exhibited different infection patterns, reflecting contrasting dynamics. Altogether, these results provide new insight into honey bee diseases and stress the need for more studies about the mechanisms of intra-colonial disease variation in social insects.

DWV Infection in vitro Using Honey Bee Pupal Tissue

The deformed wing virus (DWV) has been best characterized among honey bee viruses; however, very little is known regarding the mechanisms of viral infection and replication due to the lack of immortalized honey bee cell lines. To solve this problem, we established an in vitro system using honey bee pupal tissue to reconstruct DWV binding and entry into the host cell, followed by translation of the RNA genome and polyprotein processing using RNA-dependent RNA polymerase (RdRP) as a marker. Using this system, the P-domain of the virion subunit VP1 was found to be essential for DWV infection, but not for binding and entry into the cell. DWV efficiently infected the head tissue derived from early but not late pupa, suggesting that undifferentiated cells are targeted for viral infection. Furthermore, we found that inhibitors of mammalian picornavirus 3C-protease, rupintrivir and quercetin suppressed RdRP synthesis, indicating that this in vitro system is also useful for screening a compound to control viral infection. Our in vitro system may help to understand the mechanism of DWV infection in host cells.

Africanized honeybees in Colombia exhibit high prevalence but low level of infestation of Varroa mites and low prevalence of pathogenic viruses

The global spread of the ectoparasitic mite Varroa destructor has promoted the spread and virulence of highly infectious honey bee viruses. This phenomenon is considered the leading cause for the increased number of colony losses experienced by the mite-susceptible European honey bee populations in the Northern Hemisphere. Most of the honey bee populations in Central and South America are Africanized honey bees, which are considered more resistant to Varroa compared to European honey bees . However, the relationship between Varroa levels and spread of honey bee viruses in Africanized honey bees remains unknown. In this study, we determined Varroa prevalence and infestation levels as well as the prevalence of seven major honey bee viruses in Africanized honey bees from three regions of Colombia. We found that although Varroa exhibited high prevalence (92%), its infestation levels were low (4.6%) considering that these populations never received acaricide treatments. We also detected four viruses in the three regions analyzed, but all hives were asymptomatic, and virus prevalence was considerably lower than those found in other countries with higher rates of mite-associated colony loss ( DWV 19.88%, BQCV 17.39%, SBV 23.4 %, ABPV 10.56%). Our findings indicate that AHBs possess natural resistance to Varroa that does not prevent the spread of this parasite among their population, but restrains mite population growth and suppresses the prevalence and pathogenicity of mite-associated viruses.